Reprogramming Dermal Fibroblasts to Induced Pluripotent Stem Cells

Skin biopsies were obtained and dissected to obtain human dermal fibroblasts. They were cultured in DMEM containing L-glutamine, 15% fetal bovine serum, NEAA, and sodium pyruvate with half of the media being replaced every two days^{31 (link),91 (link)}. Once the cells were confluent, they were trypsinized and 5 × 10⁵ cells were subjected to transduction under feeder-free conditions using Yamanaka’s reprogramming factors from the CytoTune®-iPSC2.0 Sendai Reprogramming Kit, Life Technologies, A16517. When the iPSC colonies reached a size suitable for picking, they were transferred onto geltrex-coated plates^{31 (link)}. The pluripotency of the colonies was then confirmed through immunocytochemistry using TRA-1-60, SSEA4, Nanog, and OCT4 antibodies as shown in Supplementary Fig. 5.